PROJECT SUMMARY The long-term goal of this research is to elucidate the reaction mechanisms of NO detoxification catalyzed by iron-containing bacterial metalloenzymes. Our studies will combine investigations of native microbial enzymatic systems, bioengineered and synthetic models. They will focus on 1) the anaerobic reduction of NO to nitrous oxide (N2O) by denitrifying NO reductases (cNORs) and flavodiiron proteins (FDPs), and 2) the aerobic oxidation of NO to nitrate (NO3-) by members of the hemoglobin superfamily and by nonheme iron containing enzymes. All the enzymes involved in these reactions have been characterized by X-ray crystallography, but the origin of their catalytic power and whether common catalytic routes are used for each reaction remains unknown. Coupling resonance Raman, FTIR, and EPR spectroscopies with time-resolved approaches will define intermediates along these reactions. For the NO reduction reaction, we aim to define precursor to the N-N bond formation and the protonation events that must take place to convert two NO molecules to N2O and H2O. For the oxidative detoxification of NO, the major goals are to characterize iron(III)-peroxynitrite species and the mechanism of O-O bond cleavage leading to the nitrate product.